巢式PCR和实时荧光定量PCR在莱姆病宿主动物监测中的应用评价  

Evaluation of nested PCR and real-time PCR in host surveillance of Lyme disease

作　　者：张琳[1] 苗广青 侯学霞[1] 李博[2] 郝琴[1] ZHANG Lin;MIAO Guang-qing;HOU Xue-xia;LI Bo;HAO Qin(State Key Laboratory of Infectious Diseases Prevention and Control,National Institute for Communicable Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China;Xinjiang Center for Disease Control and Prevention)

机构地区：[1]中国疾病预防控制中心传染病预防控制所传染病预防控制国家重点实验室,北京102206 [2]新疆维吾尔自治区疾病预防控制中心,乌鲁木齐830001

出　　处：《中国媒介生物学及控制杂志》2018年第5期425-427,共3页Chinese Journal of Vector Biology and Control

基　　金：十三五科技重大专项(2017ZX10303404-006-003);病原监测能力建设项目(131031102000150003)~~

摘　　要：目的应用巢式PCR和实时荧光定量PCR(qRT-PCR)方法检测啮齿动物中伯氏疏螺旋体的带菌率,对2种方法在莱姆病宿主动物监测中的应用效果进行评价。方法2017年7-9月,在新疆维吾尔自治区(新疆)古尔图地区野外放置鼠夹,采集啮齿动物,并鉴别种类;采用巢式PCR和qRT-PCR 2种方法检测标本携带伯氏疏螺旋体情况,并对2种方法的检测结果进行统计学分析。对巢式PCR检测的阳性标本进行测序和序列比对分析,以初步确定伯氏疏螺旋体的基因型别。结果共检测啮齿动物标本134份,巢式PCR和qRT-PCR检测阳性率分别为13.43%和12.69%,差异无统计学意义(χ~2=0.000,P=1.000);阳性标本均为长尾黄鼠,总阳性率为20.15%,其中8份标本经2种方法检测结果均为阳性;10份巢式PCR检测为阳性的标本经qRT-PCR检测为阴性,9份qRT-PCR检测为阳性的标本经巢式PCR检测为阴性。采用q RT-PCR方法共检出伯氏疏螺旋体阳性17份,阳性样本Ct值为33.49~37.89。结论2种方法均可用于啮齿动物的伯氏疏螺旋体检测,同时使用2种方法可提高伯氏疏螺旋体的检出率。巢式PCR方法检测可以了解新疆古尔图地区啮齿动物感染伯氏疏螺旋体的基因型别;而qRT-PCR方法则可对标本中的细菌载量进行定量分析。Objective Nested PCR and quantitative real-time PCR were applied to identify the infection of Borrelia burgdorferi in rodents in this study, so that application of these two methods in host surveillance of Lyme disease can beevaluated. Methods The rodents were collected from Guertu, Xinjiang Uygur Autonomous Region during July toSeptember in 2017. The rodents were examined for the presence of B. burgdorferi by nested PCR and real-time PCR.Results In total of 134 rodent samples, the positive rate by nested PCR was 13.43% with 18 positive samples identified,whereas 17 samples were identified positive by q RT-PCR assay and the positive rate was 12.69%. The cycle threshold（Ct）value was 33.49-37.89. There was no significant difference between the two assays（ χ2=0.000, P=1.000）. All thepositives were detected from Spermophilus undulatus. Conclusion The results suggested that both nested PCR and realtime PCR could be used in identifying B. burgdorferi in rodents. Combination of these two assays could increase the positiverate. The results of nested PCR could provide local-predominant genotypes; however bacterial loads can be estimated byqRT-PCR assay.

关 键 词：伯氏疏螺旋体 啮齿动物 巢式聚合酶链式反应 实时荧光定量聚合酶链式反应 

分 类 号：R377[医药卫生—病原生物学]